1. Aspartate AminoTransferase (ASPAT)
The capacity for growth, development and yield production of a plant is influenced by the regulation of carbon and nitrogen metabolisms and the N/C ratio in a the plant Lawlor 2002 Journal of Experimental Botany, Vol. 53, No. 370, pp. 773-787.
The enzyme Aspartate aminotransferase (ASPAT enzyme) catalyzes catalyses the reversible reaction of transamination between aspartate and 2-oxoglutarate to generate glutamate and oxaloacetate using pyridoxal 5¢-phosphate (PLP) as essential cofactor in a reaction that can be express as: L-aspartate+2-oxoglutarate=oxaloacetate+L-glutamate.
The enzyme plays a key role in the metabolic regulation of carbon and nitrogen metabolism in all organisms. Structurally and functionally the ASPAT enzyme is conserved in all organisms. In eukaryots the enzyme plays a critical role in the interchanges of carbon and nitrogen pools between subcellular compartments.
Aspartate aminotransferases are classified into the group I of the aminotransferase superfamily (Jensen and Gu, 1996). Further, Aspartate Aminotransferases have been classified in four subgroups. Subgroup Ia includes the ASPATs from eubacteria and eukaryotes, whereas subgroup Ib comprises the enzymes from some eubacteria including cyanobacteria and archaebacteria. A new group of ASPAT enzymes was described by De La Torre et al. 2006 Plant J. 2006, 46(3):414-25.
In plants, genes have been identified encoding ASPAT polypeptides that are targeted to different subcellular compartments and assembled into functional ASPAT Isoenzymes in the mitochondria, the cytosol, the peroxisome and the chloroplast.
2. MYB91 Like Transcription Factor (MYB91)
DNA-binding proteins are proteins that comprise any of many DNA-binding domains and thus have a specific or general affinity to DNA. DNA-binding proteins include for example transcription factors that modulate the process of transcription, nucleases that cleave DNA molecules, and histones that are involved in DNA packaging in the cell nucleus.
Transcription factors are usually defined as proteins that show sequence-specific DNA binding affinity and that are capable of activating and/or repressing transcription. The Arabidopsis thaliana genome codes for at least 1533 transcriptional regulators, accounting for ˜5.9% of its estimated total number of genes (Riechmann et al. (2000) Science 290: 2105-2109). The Database of Rice Transcription Factors (DRTF) is a collection of known and predicted transcription factors of Oryza sativa L. ssp. indica and Oryza sativa L. ssp. japonica, and currently contains 2,025 putative transcription factors (TF) gene models in indica and 2,384 in japonica, distributed in 63 families (Gao et al. (2006) Bioinformatics 2006, 22(10):1286-7).
One of these families is the MYB domain family of transcription factors, characterized by a highly conserved DNA-binding domain, the MYB domain. The MYB domain was originally described in the oncogene (v-myb) of avian myeloblastosis virus (Klempnauer et al. (1982) Cell 33, 453-63). Many vertebrates contain three genes related to v-Myb c-Myb, A-Myb and B-Myb and other similar genes have been identified in insects, plants, fungi and slime molds. The encoded proteins are crucial to the control of proliferation and differentiation in a number of cell types. MYB proteins contain one to four imperfect direct repeats of a conserved sequence of 50-53 amino acids which encodes a helix-turn-helix structure involved in DNA binding (Rosinski and Atchley (1998) J Mol Evol 46, 74-83). Three regularly spaced tryptophan residues, which form a tryptophan cluster in the three-dimensional helix-turn-helix structure, are characteristic of a MYB repeat. The three repeats in c-Myb are referred to as R1, R2 and R3; and repeats from other MYB proteins are categorised according to their similarity to R1, R2 or R3. Since there is limited sequence conservation outside of the MYB domain, MYB proteins have been clustered into subgroups based on conserved motifs identified outside of the MYB coding region (Jiang et al. (2004) Genome Biology 5, R46).
AtMYB91 belongs to the R2R3-MYB gene family (Li and Parish, Plant J. 8, 963-972, 1995), which is a large gene family (with reportedly 126 genes in Arabidopsis thaliana (Zimmerman et al., Plant J. 40, 22-34, 2004)). Members of this group are involved in various processes, including secondary metabolism, cell morphogenesis, regulation of meristem formation, flower and seed development, cell cycle, defense and stress responses, light and hormone signalling (Chen et al., Cell Res. 16, 797-798, 2006). AtMYB91 is also named AS1 asymmetric leaves 1, and is closely related to Antirrhinum PHAN phantastica and to maize ROUGH SHEATH2 (RS2) polypeptides (Sun et al. (2002) Planta 214(5):694-702), all having an evolutionarily conserved role in specification of leaf cell identity, in particular in dorsal-ventral identity. In Arabidopsis, AS1 is expressed in leaf founder cells, where it functions as a heterodimer with the structurally unrelated AS2 proteins to repress activity of KNOTTED 1-like homeobox (KNOX) genes.
3. Gibberellic Acid-Stimulated Arabidopsis (GASA)
GASA (Gibberellic Acid-Stimulated Arabidopsis) proteins are plant-specific and are expressed during a variety of physiological processes. Several GASA-like genes are hormone responsive, expression of tomato gene GAST1, the first member of the family to be characterized, was induced upon application of exogenous gibberellin in a gibberellin-deficient background (Shi et al. Plant J. 2, 153-159, 1992). A related tomato gene, RSI-1, shares high sequence identity with GAST1 and is activated during lateral root formation (Taylor and Scheuring, Mol. Gen. Genet. 243, 148-157, 1994). GASA1 to GASA4 from Arabidopsis were first identified based on their similarity to tomato GAST1 (Herzog et al. Plant Mol. Biol. 27, 743-752, 1995). Expression data indicated that GASA1 accumulates in flower buds and immature siliques, GASA2 and GASA3 in siliques and dry seeds, and GASA4 in growing roots and flower buds. GASA4 is reported to be expressed in all meristematic regions (Aubert et al., Plant Mol. Biol. 36, 871-883, 1998).
Functionally, the GASA proteins are not well characterised. GASA proteins are reportedly involved in pathogen responses and in plant development. Plants ectopically expressing GEG, a GASA homologue from Gerbera hybrida, showed shorter corollas with decreased cell length compared with the wild type, indicating a role for GEG as an inhibitor of cell elongation. Overexpression of Arabidopsis GASA4 resulted in plants having increased seed weight (Roxrud et al, Plant Cell Physiol. 48, 471-483, 2007). However, these plants in addition had occasional meristem identity changes with reconversion from floral meristems development to normal indeterminate inflorescence development. Furthermore, modulated GASA4 expression caused a significant increase of branching. Overexpression of Arabidopsis GASA4 also increased tolerance to heat stress (Ko et al., Plant Physiol. Biochem. 45, 722-728, 2007).
4. Auxin/Indoleacetic Acid Genes (AUX/IAA)
The AUX/IAA (auxin/indoleacetic acid) genes encode a family of proteins whose expression is tightly regulated by auxin. The plant hormone auxin is involved in various processes like cell division, cell expansion and differentiation, patterning of embryos, vasculature or other tissues, regulation of growth of primary and lateral root or shoot meristems. AUX/IAA proteins furthermore are usually expressed in a tissue-specific manner.
AUX/IAA proteins typically have four conserved amino acid sequence motifs (domains I, II, III and IV) and have nuclear localisation signal sequences. Domains I and II are postulated to destabilize the protein and may be involved in protein turnover. Domains III and IV are postulated to be involved in protein-protein interactions: AUX/IAA proteins can form homodimers and are known to associate with ARF proteins. The AUX/IAA-ARF complexes are likely to be involved in auxin mediated gene expression. The Aux/IAA proteins are negative regulators of the auxin response factors (ARFs) that regulate expression of auxin-responsive genes. Aux/IAA proteins bind to the DNA-bound ARF partner proteins and repress ARF activity. In the auxin activated status, Aux/IAA proteins are ubiquitinated via interactions with the auxin-modified SCFTIR1complex and subsequently degraded by 26S proteasome action. An overview of roles and activities of AUX/IAA proteins is given by Reed (Trends in Plant Science 6, 420-425, 2001). The structure and expression analysis of early auxin-responsive Aux/IAA gene family in rice (Oryza sativa) has recently been reported by Jain et al. 2006 Funct Integr Genomics. 2006 January; 6(1):47-59.
IAA14 is a AUX/IAA protein that acts as a transcriptional repressor in lateral root formation. A gain of function mutation in IAA14 blocks early pericycle divisions that initiate lateral root development (Fukaki et al., Plant J. 29, 153-168, 2002).